The expression of CCL2 by T lymphocytes of mammary tumor bearers: role of tumor-derived factors

Tumor-associated chemokines, including CC chemokine ligand 2/monocyte chemoattractant protein-1 (CCL2), are thought to play many roles in cancer progression. Here we demonstrate the novel finding that during growth of the D1-7,12-dimethylbenzanthracene-3 mammary tumor in BALB/c mice, there is a dramatic up-regulation of CCL2 in splenic T cells at both the mRNA and protein levels upon stimulation. Of particular relevance is the finding that tumor-infiltrating T cells also produce high levels of CCL2. While a variety of tumor cell lines have been found to produce CCL2, we found no detectable levels of CCL2 protein in supernatants of the cultured mammary tumor cells. Investigation of the mechanisms involved in CCL2 induction showed that treatment of splenic T cells with the tumor-derived factors GM-CSF and phosphatidyl serine (PS) resulted in increased CCL2 production. This increased production may be involved in the downregulation of IFN-gamma by the T cells of tumor-bearing mice previously reported in this model, as treatment of splenic T lymphocytes with CCL2 resulted in a decreased secretion of IFN-gamma by those cells.     

CXCR3 in T cell function

CXCR3 is a chemokine receptor that is highly expressed on effector T cells and plays an important role in T cell trafficking and function. CXCR3 is rapidly induced on naïve cells following activation and preferentially remains highly expressed on Th1-type CD4⁺ T cells and effector CD8⁺ T cells. CXCR3 is activated by three interferon-inducible ligands CXCL9 (MIG), CXCL10 (IP-10) and CXCL11 (I-TAC). Early studies demonstrated a role for CXCR3 in the trafficking of Th1 and CD8 T cells to peripheral sites of Th1-type inflammation and the establishment of a Th1 amplification loop mediated by IFNγ and the IFNγ-inducible CXCR3 ligands. More recent studies have also suggested that CXCR3 plays a role in the migration of T cells in the microenvironment of the peripheral tissue and lymphoid compartment, facilitating the interaction of T cells with antigen presenting cells leading to the generation of effector and memory cells.     

A natural CCL5/RANTES variant antagonist for CCR1 and CCR3

The N-terminal domain of the chemokine CCL5/regulated upon activation normal T cell expressed and secreted (RANTES) has been shown to be critical for its biological activity on leukocytes. Several N-terminus-modified CCL5/RANTES derivatives, such as N-Terminal truncated CCL5/RANTES, Met-RANTES, and amino-oxypentane (AOP)-RANTES exhibited antagonist or partial agonist functions when investigated on the properties of their receptors CCR1, CCR3, and CCR5. Studying 95 African samples from Cameroon, we found a naturally occurring variant of CCL5/RANTES containing a missense mutation located in the first amino acid of the secreted form (S24F). S24F binds CCR1, CCR3, and CCR5 and triggers receptor down-modulation comparable to CCL5/RANTES. Moreover, in CCR5 positive cells, S24F elicits cellular calcium mobilization equivalent to that obtained with CCL5/RANTES. By contrast, S24F does not provoke any response in CCR1 and CCR3 positive cells. As CCL5/RANTES is able to attract different subtypes of leukocytes into inflamed tissue and intervenes in a wide range of allergic and autoimmune diseases, the discovery of this natural N-terminus-modified CCL5/RANTES analogue exhibiting differential effects on CCL5/RANTES receptors, opens up additional perspectives for therapeutic intervention.Nucleotide sequence data reported is available in the DDBJ/EMBL/GenBank databases under the accession number: DQ230537.     

CXCR5 may be involved in the attraction of human metastatic neuroblastoma cells to the bone marrow

Introduction Up-regulation of some chemokine receptors on tumor cells is associated with increased metastatic potential. In this respect, limited information is available on chemokine receptor in human neuroblastoma (NB). Objects Purpose of the study was to identify chemokines/chemokine receptors involved in bone marrow (BM) localization of metastatic NB cells in view of the development of targeted therapeutic strategies. CD45⁻ metastatic NB cells were isolated from the BM of six patients by immunomagnetic bead manipulation. Some experiments were carried out using a panel of human neuroblastoma cell lines (GI-ME-N, GI-LI-N, LAN-5, HTLA-230, SH-SY-5Y and IMR-32). Immunophenotypic analyses were performed by flow cytometry. Cell migration assays were carried out using transwell systems. Calcium ion mobilization, chemokine receptor internalization and cell proliferation were investigated by flow cytometry. Results In all BM samples, CXCR5 was expressed by the majority of primary neuroblasts and mediated their chemotaxis in response to CXCL13. Primary metastatic NB cells from all BM samples expressed CXCR6, but were not attracted by soluble CXCL16. Studies performed with two CXCR6⁺ NB cell lines showed that the mechanism whereby neuroblasts did not migrate to CXCL16 was likely related to defective calcium ion mobilization. Conclusions CXCR5 is the first chemokine receptor so far identified able to attract in vitro primary metastatic NB cells. CXCR6 may be involved in retention of metastatic neuroblasts in the BM through interaction with CXCL16 expressing stromal cells in the absence of signal transduction.     

Expression of CXC and CC type of chemokines and its receptors in tuberculous and non-tuberculous effusions

Chemokines mediate their biological functions by transmigration of various immune cells to the site of infection. Tuberculous pleurisy provides an effective model to study the role of chemokines in the recruitment of immune cells to the pleura. Our aim was to understand the cumulative effect of chemokines (IP-10, MIG, IL-8, MCP-1, MIP-1α and RANTES) and its receptors (CXCR2, CXCR3, CCR1, CCR2, CCR5 and CCR7) in the recruitment of CD4⁺ T cells obtained from blood (BL) and pleural fluid (PF) of tuberculous (TB) and non-tuberculous (NTB) patients. We observed significant increase in CD4⁺ T cells in TB PF indicating lymphocytic rich effusion. All chemokines except RANTES were significantly high in PF compared to BL in TB group, whereas IL-8 and MCP-1 showed significant increase only in NTB PF. The significantly high levels of IFN-γ and ΤΝF-α in TB PF and their positive correlation with IP-10 and MIP-1α indicated their synergistic action to elicit a strong protective Th1 response. In spite of high levels of Th1 cytokines and chemokines in TB PF, significantly lower levels of RANTES indicated its limited role at the site. The CXC receptors in PF of both the groups and CC receptors except CCR5 in TB PF were significantly high compared to BL. Only CXCR2, CCR5 and CCR7 showed significant increase in TB compared to NTB. Thus a selective concentration of chemokines, cytokines and abundant expression of chemokine receptors confirm the accumulation of activated and memory T cells at the site of infection and help in polarizing Th1 immune response.     

趋化因子及趋化因子受体3在自身免疫疾病中的作用

人体在防御和清除入侵病原体等异物时,有一种使白细胞趋集的功能,有一些低分子量的物质能引起这种功能称之为趋化剂或趋化因子。这些小蛋白因其有定向细胞趋化作用而得名。经研究表明,趋化因子受体3（CXCR3）趋化因子可能在自身免疫内分泌疾病中起到致病作用。此外,血清中CXCR3趋化因子的判定可能辅助检测免疫活性。CXCR3和优先参与趋化Th1细胞的因子。该受体连接的趋化因子10（CXCL10）不仅参与白细胞募集,还诱导T细胞增殖的异源体和抗原的刺激。趋化因子10正调节Th1细胞产物并且负调节Th2细胞的产物。免疫反应纤维结合素（INF）产物可增强特异的炎症反应。当被激活或者发现炎症和肿瘤细胞后趋化因子受体3-B在内皮细胞中表达并且其结合的趋化因子10,趋化因子9和趋化因子11激活后产生血管抑制作用。     

Expression of the inflammatory chemokines CCL5, CCL3 and CXCL10 in juvenile idiopathic arthritis, and demonstration of CCL5 production by an atypical subset of CD8+ T cells

This study focuses upon three chemokines, namely CCL5, CXCL10 and CCL3, which are potential novel therapeutic targets in arthritis. The aim of the study was to analyse the expression and production of these three chemokines within the joints of children with juvenile idiopathic arthritis (JIA) of the oligoarticular and polyarticular subtypes. All three of these chemokines are highly expressed at the level of mRNA, with the most significant increase in mRNA levels being demonstrated for CCL5 when compared with matched peripheral blood samples and controls. We show that high levels of all three chemokines are present in synovial fluid of children with JIA. We investigate the major source of CCL5 from inflammatory synovial cells, which we show to be CD8+ T cells. This CD8+ synovial T cell population has an unexpected phenotype that has not been described previously, being CCR7- yet predominantly CD28+ and CD45RA-. These cells contain high levels of stored intracellular CCL5, and rapid release of CCL5 takes place on T cell stimulation, without requiring new protein synthesis. In addition, we demonstrate that CCL5 is present in synovial biopsies from these patients, in particular on the endothelium of small and medium sized vessels. We believe this to be the first in depth analysis of these mediators of inflammation in JIA.     

Towards in situ tissue repair: human mesenchymal stem cells express chemokine receptors CXCR1, CXCR2 and CCR2, and migrate upon stimulation with CXCL8 but not CCL2

The recruitment of bone marrow CD34- mesenchymal stem- and progenitor cells (MSC) and their subsequent differentiation into distinct tissues is the precondition for in situ tissue engineering. The objective of this study was to determine the entire chemokine receptor expression profile of human MSC and to investigate their chemotactic response to the selected chemokines CCL2, CXCL8 and CXCL12. Human MSC were isolated from iliac crest bone marrow aspirates and showed a homogeneous population presenting a typical MSC-related cell surface antigen profile (CD14-, CD34-, CD44+, CD45-, CD166+, SH-2+). The expression profile of all 18 chemokine receptors was determined by real-time PCR and immunohistochemistry. Both methods consistently demonstrated that MSC express CC, CXC, C and CX(3)C receptors. Gene expression and immunohistochemical analysis documented that MSC express chemokine receptors CCR2, CCR8, CXCR1, CXCR2 and CXCR3. A dose-dependent chemotactic activity of CXCR4 and CXCR1/CXCR2 ligands CXCL12 and CXCL8 (interleukin-8) was demonstrated using a 96-well chemotaxis assay. In contrast, the CCR2 ligand CCL2 (monocyte chemoattractant protein-1, MCP-1) did not recruited human MSC. In conclusion, we report that the chemokine receptor expression profile of human MSC is much broader than known before. Furthermore, for the first time, we demonstrate that human MSC migrate upon stimulation with CXCL8 but not CCL2. In combination with already known data on MSC recruitment and differentiation these are promising results towards in situ regenerative medicine approaches based on guiding of MSC to sites of degenerated tissues.     

Analysis of NK cells and chemokine receptors in tumor infiltrating CD4 T lymphocytes in human renal carcinomas

Recent data suggest that chemokines and chemokine receptors mediate leukocyte recruitment of all components of the antitumor response. This study aimed to phenotypically characterize the immune lymphocyte infiltrate in human renal cell carcinomas (RCCs) and at the invasive margin (tumor–host interface) and to define the association of these findings with established prognostic indicators. Tumor infiltrating lymphocytes (TILs) were obtained from 24 patients with RCC undergoing radical nephrectomy. Peripheral blood cells from 37 patients were also obtained before surgery. Our findings are consistent with the preferential recruitment of CD4+ Th1-polarized effector memory cells that express CXCR3/CCR5. These cells were the main component of TILs and expressed as CXCR3, CCR5, CD45RO, and CD95. Natural killer (NK) cells were found in significantly higher proportions in TILs of RCCs than in peripheral blood lymphocytes (PBLs) or in other tumors studied (colorectal and breast cancers), where these cells were found in small proportions. No differences in nuclear grade or other studied parameters were observed between the TILs and the lymphocytes present at the invasive margin, which showed a similar composition. However, differences were found according to the tumor stage. First, significantly fewer NK cells were observed in PBLs from metastatic patients. Second, a significantly lower proportion of CCR5/CXCR3/CD4+ cells and a higher proportion of CCR4/CD4+ cells were observed in metastatic patients, suggesting that preferential Th1-polarization may gradually change during the progression of renal cancer cells. Finally, the frequency of CD25/CD4+ cells was higher in metastatic patients. Although the sample of patients with metastasis was small, the overall results suggest a change in composition of the TILs that may potentially confer a selective advantage for tumor growth and may account for the suppression of an effective cytotoxic response.     

CCR7/CCL21 migration on fibronectin is mediated by phospholipase Cgamma1 and ERK1/2 in primary T lymphocytes

CCR7 binds to its cognate ligand, CCL21, to mediate the migration of circulating naive T lymphocytes to the lymph nodes. T lymphocytes can bind to fibronectin, a constituent of lymph nodes, via their β1 integrins, which is a primary mechanism of T lymphocyte migration; however, the signaling pathways involved are unclear. We report that rapid (within 2 min) and transient phosphorylation of ERK1/2 is required for T cell migration on fibronectin in response to CCL21. Conversely, prevention of ERK1/2 phosphorylation by inhibition of its kinase, MAPK/MEK, prevented T lymphocyte migration. Previous studies have suggested that phospholipase Cγ1 (PLCγ1) can mediate phosphorylation of ERK1/2, which is required for β1 integrin activation. Paradoxically, we found that inhibition of PLCγ1 phosphorylation by the general PLC inhibitor U73122 was associated with a delayed and reduced phosphorylation of ERK1/2 and reduced migration of T lymphocytes on fibronectin. To further characterize the relationship between ERK1/2 and PLCγ1, we reduced PLCγ1 levels by 85% using shRNA and observed a reduced phosphorylation of ERK1/2 and a significant loss of CCR7-mediated migration of T lymphocytes on fibronectin. In addition, we found that inhibition of ERK1/2 phosphorylation by U0126 resulted in a decreased phosphorylation of PLCγ1, suggesting a feedback loop between ERK1/2 and PLCγ1. Overall, these results suggest that the CCR7 signaling pathway leading to T lymphocyte migration on fibronectin is a β1 integrin-dependent pathway involving transient ERK1/2 phosphorylation, which is modulated by PLCγ1.     

Colon carcinoma cells induce CXCL11-dependent migration of CXCR3-expressing cytotoxic T lymphocytes in organotypic culture

Adoptive immunotherapy of cancer patients with cytolytic T lymphocytes (CTL) has been hampered by the inability of the CTL to home into tumors in vivo. Chemokines can attract T lymphocytes to the tumor site, as demonstrated in animal models, but the role of chemokines in T-lymphocyte trafficking toward human tumor cells is relatively unexplored. In the present study, the role of chemokines and their receptors in the migration of a colon carcinoma (CC) patient’s CTL toward autologous tumor cells has been studied in a novel three-dimensional organotypic CC culture. CTL migration was mediated by chemokine receptor CXCR3 expressed by the CTL and CXCL11 chemokine secreted by the tumor cells. Excess CXCL11 or antibodies to CXCL11 or CXCR3 inhibited migration of CTL to tumor cells. T cell and tumor cell analyses for CXCR3 and CXCL11 expression, respectively, in ten additional CC samples, may suggest their involvement in other CC patients. Our studies, together with previous studies indicating angiostatic activity of CXCL11, suggest that CXCL11 may be useful as an immunotherapeutic agent for cancer patients when transduced into tumor cells or fused to tumor antigen-specific Ab.     

CXCL8((3-73))K11R/G31P antagonizes ligand binding to the neutrophil CXCR1 and CXCR2 receptors and cellular responses to CXCL8/IL-8

We recently reported that CXCL8((3-73))K11R is a high affinity agonist of neutrophil activation and chemotactic responses. In this report we employed CXCL8((3-73))K11R as a template to generate CXCL8/IL-8 analogues with antagonist activities, using site-directed mutagenesis to introduce conservative amino acid substitutions into the first turn within the molecule's beta-pleated sheet region (G31P, P32G) and, in association with these, into the putative receptor-recognition site (T12S, H13F, F17S). We then examined their impact on the analogues' biological activities and found that a G31P substitution rendered CXCL8((3-73))K11R a high affinity antagonist of CXCL8/IL-8. The ranking (in the order of decreasing CXCL8/IL-8 antagonist activities) of the CXCL8((3-73))K11R analogues we generated was, G31P>T12S/G31P>H13F/G31P>T12S/H13F/G31P>P32G approximately T12S/P32G approximately H13F/P32G>T12S/H13F/P32G; CXCL8((3-73))K11R/F17S did not inhibit CXCL8/IL-8-dependent responses. CXCL8((3-73))K11R/G31P had no discernible agonist (beta-glucuronidase release, chemotactic) activity, but at 12.5 ng/ml it bound to purified neutrophils more avidly than did 1.25 microg/ml CXCL8/IL-8. Furthermore, CXCL8((3-73))K11R/G31P competitively antagonized the binding of CXCR1- and CXCR2-specific antibodies to these receptors. Taken together, these data thus provide further impetus to the study of the potential efficacy of CXCL8((3-73))K11R/G31P as a broad-spectrum antagonist of the ELR-CXC chemokines in experimental and clinical settings.     

Cleavage of chemokines CCL2 and CXCL10 by matrix metalloproteinases-2 and -9: Implications for chemotaxis

Proteolytic processing of chemokines is a complex process that can result in dramatic effects on their chemotactic activity. Results from gel electrophoresis and mass spectrometry using recombinant CCL2 and CXCL10, incubated with either MMP-2 or -9, indicate that both chemokines are cleaved by the enzymes. N-terminal truncation of four amino acids from CCL2, and four or five residues from CXCL10 occurred, but removal of four residues from the C-terminus of CXCL10 was also observed with both MMPs. The speed of the reaction was chemokine-dependent, with N-terminal processing of CCL2 being complete within 3 h, whereas activity of the MMPs on CXCL10 remained incomplete at 48 h. The effect on the chemotactic potential of N-terminal truncation of CCL2 by MMPs-2 and -9 was investigated using in vitro migration assays. Monocytic cells exhibited a 2-fold reduction in migration to MMP-cleaved CCL2 variants, compared to intact CCL2.     

Muscarinic receptors are involved in LMM3 tumor cells proliferation and angiogenesis

Angiogenesis is a process of new blood vessel development from pre-existing vasculature and it plays an essential role in tumor growth and metastases. Here, we investigate the expression of muscarinic acetylcholine receptors (mAchR) and their participation in tumor cell proliferation and angiogenesis ability. Saturation binding assays with the tritiated muscarinic antagonist quinuclidinyl benzilate indicate that LMM3 cells derived from a murine mammary adenocarcinoma express a single class of functional mAchR. Competition binding assays with selective muscarinic antagonists indicate a predominance of M3 receptor subtype. The muscarinic agonist carbachol (CARB) stimulates LMM3 cell proliferation in a concentration dependent manner. The maximal effect induced by 10(-9)M CARB was totally blunted by atropine and by the selective M3 and M1 antagonists, para-fluoro hexahydro sila-difenidol (pf-HHSiD) and pirenzepine, respectively. In addition, pf-HHSiD completely blocked in vivo CARB-induced neovascular formation and vascular endothelial growth factor-A in LMM3 tumor cells. We can conclude that mAchR expressed in LMM3 mammary tumor cells positively regulate proliferation and angiogenesis required for tumor progression.     

Expression and characterization of the chemokine receptors CCR2 and CCR5 in mice

The chemokine receptors CCR2 and CCR5 play important roles in the recruitment of monocytes/macrophages and T cells. To better understand the role of both receptors in murine models of inflammatory diseases and to recognize potential problems when correlating these data to humans, we have generated mAbs against murine CCR2 and CCR5. In mice CCR2 is homogeneously expressed on monocytes and on 2--15% of T cells, closely resembling the expression pattern in humans. In contrast to humans, murine NK cells are highly CCR5 positive. In addition, CCR5 is expressed on 3--10% of CD4 and 10--40% of CD8-positive T cells and is weakly detectable on monocytes. Using a model of immune complex nephritis, we examined the effects of inflammation on chemokine receptor expression and found a 10-fold enrichment of CCR5(+) and CCR2(+) T cells in the inflamed kidneys. The activity of various chemokines and the antagonistic properties of the mAbs were measured by ligand-induced internalization of CCR2 and CCR5 on primary leukocytes. The Ab MC-21 (anti-CCR2) reduced the activity of murine monocyte chemotactic protein 1 by 95%, whereas the Ab MC-68 (anti-CCR5) blocked over 99% of the macrophage-inflammatory protein 1alpha and RANTES activity. MC-21 and MC-68 efficiently blocked the ligand binding to CCR2 and CCR5 with an IC(50) of 0.09 and 0.6--1.0 microg/ml, respectively. In good correlation to these in vitro data, MC-21 almost completely prevented the influx of monocytes in thioglycollate-induced peritonitis. Therefore, both Abs appear as useful reagents to further study the role of CCR2 and CCR5 in murine disease models.     

Higher circulating levels of chemokines CXCL10, CCL20 and CCL22 in patients with ischemic heart disease

Recruitment of leukocytes is one of the earliest events in the pathogenesis of ischemic heart disease (IHD) and chemokines play an important role in the migration of these cells into the inflammation sites. The aim of this study was to evaluate the CXCL10, CCL20 and CCL22 levels and the single nucleotide polymorphisms (SNPs) rs4508917, rs6749704 and rs4359426 in chemokine genes in patients with IHD to clarify any association. A total of 300 patients with IHD as having acute myocardial infarction (AMI; n = 100), stable angina (SA; n = 100) or unstable angina (UA; n = 100) and 100 healthy subjects as a control group were enrolled to study. Serum samples from all participants were tested for the CXCL10, CCL20 and CCL22 levels by using ELISA. The SNPs were determined by polymerase chain reaction–restriction length polymorphism (PCR–RFLP) method. The mean serum concentrations of CXCL10, CCL20 and CCL22 in AMI patients (395.97 ± 21.20 Pg/mL, 108.38 ± 10.31 Pg/mL and 1852.58 ± 205.77 Pg/mL), SA patients (405.48 ± 27.36 Pg/mL, 90.20 ± 7.69 Pg/mL and 2322.04 ± 231.23 Pg/mL) and UA patients (396.69 ± 22.79 Pg/mL, 141.87 ± 18.10 Pg/mL and 2754.89 ± 211.70 Pg/mL) were significantly higher than in the healthy group (179.38 ± 8.85 Pg/mL, 51.92 ± 4.62 Pg/mL and 451.82 ± 23.76 Pg/mL, respectively; P < 0.001). Similarly, the serum levels of CXCL10, CCL20 and CCL22 in total IHD patients (399.38 ± 13.77 Pg/mL, 113.49 ± 7.48 Pg/mL and 2309.84 ± 126.39 Pg/mL, respectively) were also significantly higher as compared with healthy subjects (P < 0.001). The serum levels of CCL20 and CCL22 in UA patients were significantly higher than those in SA and AMI patients, respectively (P < 0.01 and P < 0.003, respectively). The serum levels of CXCL10 and CCL20 in diabetic patients were significantly higher in comparison to non-diabetic patients (P < 0.05 and P < 0.02, respectively). The serum levels of CCL22 in dyslipidemic- and obese patients were also significantly higher in comparison with non-dyslipidemic- and non-obese patients, correspondingly (P < 0.05 and P < 0.01, respectively). There were no significant differences between men and women or between patients who treated with statin, aspirin, β-blockers or angiotensin converting enzyme (ACE) inhibitors and patients without mentioned treatment regarding the levels of chemokines. The frequency of the GG genotype at SNP rs4508917 in CXCL10 gene was higher, whereas the frequency of the AA genotype at SNP rs4359426 in CCL22 gene was lower in total patients with IHD as compared with healthy subjects (P < 0.04 and P < 0.002, respectively). These results showed that the higher levels of CXCL10, CCL20 and CCL22 were associated with IHD. The serum levels of chemokines may influence by the certain traditional risk factors of IHD and some studied SNPs, but did not influence by treatment and gender of patients.     

Protein kinase D1 and D2 are involved in chemokine release induced by toll-like receptors 2, 4, and 5

The protein kinase D (PKD) family consists of three serine-threonine kinases involved in cellular proliferation, motility, and apoptosis. We previously reported that human toll-like receptor 5 (TLR5) contains a consensus PKD phosphorylation site. Flagellin stimulation of cells activated PKD1, and inhibition of PKD1 reduced flagellin-induced interleukin-8 (IL-8) production in epithelial cells. In the current work, we examined PKD1 and PKD2 involvement downstream of TLR5, TLR4 and TLR2. We found that inhibition of either kinase with shRNA reduced IL-8 and CCL20 release due to TLR4 and TLR2 agonists to a similar extent as previously reported for TLR5. PKD1 and PKD2 inhibition reduced NF-κB activity but not MAPK activation. These results demonstrate that both PKD1 and PKD2 are required for inflammatory responses following TLR2, TLR4, or TLR5 activation, although PKD1 is more strongly involved. These kinases likely act downstream of the TLRs themselves to facilitate NF-κB activation but not MAP kinase phosphorylation.     

gp120 induces cell death in human neuroblastoma cells through the CXCR4 and CCR5 chemokine receptors

To infect target cells, the human immunodeficiency virus (HIV) type I (HIV-1) must engage not only the well-known CD4 molecule, but it also requires one of several recently described coreceptors. In particular, the CXCR4 (LESTR/fusin) receptor allows fusion and entry of T-tropic strains of HIV, whereas CCR5 is the major coreceptor used by primary HIV-1 strains that infect macrophages and CD4(+) T-helper cells (M-tropic viruses). In addition, the alpha chemokine SDF1alpha and the beta chemokines MIP1alpha, MIP1beta, and RANTES, natural ligands of CXCR4 and CCR5, respectively, are potent soluble inhibitors of HIV infection by blocking the binding between the viral envelope glycoprotein gp120 and the coreceptors. Approximately two-thirds of individuals with acquired immunodeficiency syndrome (AIDS) show neurologic complications, which are referred to a syndrome called AIDS dementia complex or HIV-1-associated cognitive/motor complex. The HIV-1 coat glycoprotein gp120 has been proposed as the major etiologic agent for neuronal damage, mediating both direct and indirect effects on the CNS. Furthermore, recent findings showing the presence of chemokine receptors on the surface of different cell types resident in the CNS raise the possibility that the association of gp120 with these receptors may contribute to the pathogenesis of neurological dysfunction. Here, we address the possible role of alpha and beta chemokines in inhibiting gp120-mediated neurotoxicity using the human neuroblastoma CHP100 cell line as an experimental model. We have previously shown that, in CHP100 cells, picomolar concentrations of gp120 produce a significant increase in cell death, which seems to proceed through a Ca(2+) - and NMDA receptor-dependent cascade. In this study, we gained insight into the mechanism(s) of neurotoxicity elicited by the viral glycoprotein. We found that CHP100 cells constitutively express both CXCR4 and CCR5 receptors and that stimulation with phorbol 12-myristate 13-acetate down-regulates their expression, thus preventing gp120-induced cell death. Furthermore, all the natural ligands of these receptors exerted protective effects against gp120-mediated neuronal damage, although with different efficiencies. These findings, together with our previous reports, suggest that the neuronal injury observed in HIV-1 infection could be due to direct (or indirect) interactions between the viral protein gp120 and chemokine and/or NMDA receptors.     

Higher expression of CCL2, CCL4, CCL5, CCL21, and CXCL8 chemokines in the skin associated with parasite density in canine visceral leishmaniasis

Background

The immune response in the skin of dogs infected with Leishmania infantum is poorly understood, and limited studies have described the immunopathological profile with regard to distinct levels of tissue parasitism and the clinical progression of canine visceral leishmaniasis (CVL).

Methodology/Principal Findings

A detailed analysis of inflammatory cells (neutrophils, eosinophils, mast cells, lymphocytes, and macrophages) as well as the expression of chemokines (CCL2, CCL4, CCL5, CCL13, CCL17, CCL21, CCL24, and CXCL8) was carried out in dermis skin samples from 35 dogs that were naturally infected with L. infantum. The analysis was based on real-time polymerase chain reaction (PCR) in the context of skin parasitism and the clinical status of CVL. We demonstrated increased inflammatory infiltrate composed mainly of mononuclear cells in the skin of animals with severe forms of CVL and high parasite density. Analysis of the inflammatory cell profile of the skin revealed an increase in the number of macrophages and reductions in lymphocytes, eosinophils, and mast cells that correlated with clinical progression of the disease. Additionally, enhanced parasite density was correlated with an increase in macrophages and decreases in eosinophils and mast cells. The chemokine mRNA expression demonstrated that enhanced parasite density was positively correlated with the expression of CCL2, CCL4, CCL5, CCL21, and CXCL8. In contrast, there was a negative correlation between parasite density and CCL24 expression.

Conclusions/Significance

These findings represent an advance in the knowledge about skin inflammatory infiltrates in CVL and the systemic consequences. Additionally, the findings may contribute to the design of new and more efficient prophylactic tools and immunological therapies against CVL.     

Regulation of activities of NK cells and CD4 expression in T cells by human HNP-1, -2, and -3

Human neutrophil defensin alpha (HNP) is a group of cationic peptides of diverse physiological roles. Recent studies revealed the nature of HNPs as the dominant HLA-DR binding peptides on malignant cancer cells, which may block the major histocompatibility complex for antigen presentation. Here we show that HNPs may inhibit T cells by downregulating CD4 expression, a molecule of critical importance for T cell's interaction with the target cell. HNPs also inhibited tumor-cell-lysis activities of NK cells by downregulating CD16-CD56 expression. More importantly, HNPs were markedly elevated in 14 cancer tissues out of 15 self-paired human colorectal cancers and their adjacent noncancerous tissues. The subset compositions of HNPs extracted from cancer tissues and neutrophils were identical. Immunohistochemical studies indicated that HNPs mainly distributed in the infiltrated neutrophils in the interstitium. The elevated HNPs in cancer tissues may create a microenvironment unfavorable for adaptive immune reaction, implicating the cancer evasion.